title: G protein-dependent basal and evoked endothelial cell vWF secretion.
abstract: von Willebrand factor (vWF) secretion by endothelial cells (ECs) is essential for hemostasis and thrombosis; however, the molecular mechanisms are poorly understood. Interestingly, we observed increased bleeding in EC-Gα13(-/-);Gα12(-/-) mice that could be normalized by infusion of human vWF. Blood from Gα12(-/-) mice exhibited significantly reduced vWF levels but normal vWF multimers and impaired laser-induced thrombus formation, indicating that Gα12 plays a prominent role in EC vWF secretion required for hemostasis and thrombosis. In isolated buffer-perfused mouse lungs, basal vWF levels were significantly reduced in Gα12(-/-), whereas thrombin-induced vWF secretion was defective in both EC-Gαq(-/-);Gα11(-/-) and Gα12(-/-) mice. Using siRNA in cultured human umbilical vein ECs and human pulmonary artery ECs, depletion of Gα12 and soluble N-ethylmaleimide-sensitive-fusion factor attachment protein α (α-SNAP), but not Gα13, inhibited both basal and thrombin-induced vWF secretion, whereas overexpression of activated Gα12 promoted vWF secretion. In Gαq, p115 RhoGEF, and RhoA-depleted human umbilical vein ECs, thrombin-induced vWF secretion was reduced by 40%, whereas basal secretion was unchanged. Finally, in vitro binding assays revealed that Gα12 N-terminal residues 10-15 mediated the binding of Gα12 to α-SNAP, and an engineered α-SNAP binding-domain minigene peptide blocked basal and evoked vWF secretion. Discovery of obligatory and complementary roles of Gα12 and Gαq/11 in basal vs evoked EC vWF secretion may provide promising new therapeutic strategies for treatment of thrombotic disease.
authors.affiliation: Department of Pharmacology and.
authors.forename: Luiza
authors.initials: L
authors.lastname: Rusu
authors.forename: Alexandra
authors.initials: A
authors.lastname: Andreeva
authors.forename: David J
authors.initials: DJ
authors.lastname: Visintine
authors.forename: Kyungho
authors.initials: K
authors.lastname: Kim
authors.forename: Stephen M
authors.initials: SM
authors.lastname: Vogel
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Stojanovic-Terpo
authors.forename: Olga
authors.initials: O
authors.lastname: Chernaya
authors.forename: Guoquan
authors.initials: G
authors.lastname: Liu
authors.forename: Farnaz R
authors.initials: FR
authors.lastname: Bakhshi
authors.forename: Sandra L
authors.initials: SL
authors.lastname: Haberichter
authors.forename: Hiroko
authors.initials: H
authors.lastname: Iwanari
authors.forename: Osamu
authors.initials: O
authors.lastname: Kusano-Arai
authors.forename: Nobuchika
authors.initials: N
authors.lastname: Suzuki
authors.forename: Takao
authors.initials: T
authors.lastname: Hamakubo
authors.forename: Tohru
authors.initials: T
authors.lastname: Kozasa
authors.forename: Jaehyung
authors.initials: J
authors.lastname: Cho
authors.forename: Xiaoping
authors.initials: X
authors.lastname: Du
authors.forename: Richard D
authors.initials: RD
authors.lastname: Minshall
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry; D042783:Endothelial Cells / Q000166:cytology*; D044365:GTP-Binding Protein alpha Subunits, G12-G13 / Q000378:metabolism*; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000378:metabolism*; D005786:Gene Expression Regulation; D006487:Hemostasis; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D010973:Platelet Adhesiveness; D011485:Protein Binding; D034741:RNA, Small Interfering / Q000378:metabolism; D015398:Signal Transduction; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins / Q000737:chemistry; D013927:Thrombosis; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*; D014841:von Willebrand Factor / Q000378:metabolism*
chemical_list: D000911:Antibodies, Monoclonal; D034741:RNA, Small Interfering; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D014841:von Willebrand Factor; D044365:GTP-Binding Protein alpha Subunits, G12-G13; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D020742:rhoA GTP-Binding Protein